腺苷一磷酸激活蛋白激酶激活劑研究進展

徐亞洲，廖紅，張陸勇，李佳，龐濤*

（1. 中國藥科大學新藥篩選中心，江蘇 南京 210009；2. 中國科學院上海藥物研究所，國家新藥篩選中心，上海 201203）

腺苷一磷酸激活蛋白激酶激活劑研究進展

徐亞洲1，廖紅1，張陸勇1，李佳2，龐濤1*

（1. 中國藥科大學新藥篩選中心，江蘇 南京 210009；2. 中國科學院上海藥物研究所，國家新藥篩選中心，上海 201203）

腺苷一磷酸激活蛋白激酶（AMPK）是調(diào)控能量代謝的重要激酶，在代謝障礙、心血管疾病及腫瘤等疾病的病理進程中都有重要的調(diào)節(jié)作用。對AMPK的結(jié)構(gòu)及其生理調(diào)節(jié)作用進行介紹，并重點綜述AMPK間接激活劑和直接激活劑的研究進展，旨在為AMPK激活劑的深入開發(fā)提供參考。

腺苷一磷酸激活蛋白激酶；間接激活劑；直接激活劑

腺苷一磷酸激活蛋白激酶（adenosine 5'-monophosphateactivated protein kinase, AMPK）廣泛存在于生物體內(nèi)，對細胞能量代謝、生長分化等發(fā)揮關(guān)鍵作用，是一種極其重要的蛋白激酶，其生物學效應及其激活劑在臨床疾病治療方面的研究日益廣泛。目前認為，AMPK作為靶點在治療心血管疾病、糖尿病[1]、肥胖癥[2]、中樞性疾病[3]以及癌癥[4]等方面有著很大的應用潛力。

1 AMPK的結(jié)構(gòu)

AMPK屬絲氨酸/蘇氨酸蛋白激酶，是由α、β和γ3個亞基組成的異源三聚體復合物（見圖1）。其中，α亞基為催化亞基，有α1和α2這2種亞型；β、γ亞基為調(diào)節(jié)亞基，有β1、β2、γ1、γ2和γ3亞型，各種亞型由獨立的基因編碼，可組成多種可能的復合體形式[5]。這些復合體在機體不同組織中有不同的表達，且有可能發(fā)揮不同的作用，其中α2、β2在心臟和骨骼肌中高表達，γ2在心臟、腦等少數(shù)組織中表達，γ3只在骨骼肌中表達，而α1、β1、γ1在機體各組織中普遍存在[6]。最新研究表明：人類骨骼肌中AMPK復合體主要為α2β2γ3型[7]，肝臟內(nèi)的AMPK復合體主要為α1β2γ1型[8]。

AMPK的α亞基N端含1個高度保守的催化結(jié)構(gòu)域，其172位蘇氨酸（Thr172）為磷酸化激活位點，另外還包括1個自抑制區(qū)（auto-inhibitory domain，AID）以及C端1個與β亞基和γ亞基結(jié)合的區(qū)域。β亞基包含1個N端豆蔻?；稽c(N-terminal myristoylation site)、中間的糖原結(jié)合區(qū)域和C端1個與其他2個亞基的結(jié)合區(qū)。γ亞基4個胱硫醚-β-合成酶（cystathionine-β-synthase，CBS）序列，組成2個Bateman域，每個Bateman域能結(jié)合1個腺嘌呤核苷酸如AMP或ATP[6-7]。

圖1 腺苷一磷酸激活蛋白激酶結(jié)構(gòu)圖Figure 1 Structure of AMPK

2 AMPK的生理調(diào)節(jié)

目前認為，體內(nèi)存在多種激酶可激活AMPK（見圖2），主要有2種：一是肝激酶B1（liver kinase B1，LKB1），依賴細胞內(nèi)AMP激活AMPK；二是鈣/鈣調(diào)蛋白依賴蛋白激酶激酶β（calcium/calmodulin-dependent protein kinase kinase beta，CaMKKβ），依賴第2信使鈣離子的參與[9]。LKB1存在于除Hela細胞外的大多數(shù)組織細胞內(nèi)，而CaMKKβ在中樞系統(tǒng)中高度表達且發(fā)揮著重要作用[6,10]。此外，還存在轉(zhuǎn)化生長因子β激活激酶（transforming growth factor beta-activated kinase-1，Tak1）能激活AMPK。這3種激酶都作用于α亞基上的Thr172位點，使其磷酸化進而激活AMPK[7]。體內(nèi)的蛋白磷酸酶2C（protein phosphotases 2C，PP2C）可催化AMPK的Thr172位點的去磷酸化[11]，使其變成無活性形式。

生理狀態(tài)下，單磷酸腺苷（adenosine monophosphate，AMP）結(jié)合到γ亞基Bateman域上變構(gòu)激活AMPK，同時AMP與γ亞基的結(jié)合促進α亞基上Thr172的磷酸化并阻礙PP2C的去磷酸化效應，從而大大增強了AMPK的活性[6,11]。研究表明，ATP可拮抗AMP結(jié)合到γ亞基上[12]，因此，生理或病理狀態(tài)下只要能引起細胞內(nèi)AMP/ATP比值細微變化的因素均能影響AMPK活性，如運動、激素刺激、缺血、缺氧、氧化應激等。激活的AMPK可激活其下游通路，從而發(fā)揮一系列重要作用，如調(diào)控營養(yǎng)物質(zhì)代謝[1]、抑制促炎因子的合成釋放[13]、影響腫瘤細胞生長[4]等。因此，AMPK已成為廣泛關(guān)注的多種疾病的治療新靶點，其激活劑也成為藥物研究與開發(fā)的熱點之一。

圖2 腺苷一磷酸激活蛋白激酶的生理調(diào)節(jié)Figure 2 Physiological regulation of AMPK

3 AMPK激活劑

3.1 間接激活劑

很多藥物和激素可在體內(nèi)激活AMPK，但由于這些激活劑的作用機制尚未完全闡釋清楚，且目前的研究證明其并不能與AMPK直接作用，而是通過影響AMP/ATP比值或AMPK上游激酶活性等途徑間接激活AMPK，此類激活劑被稱為間接激活劑（部分間接激活劑的作用機制見圖3）。

圖3 腺苷一磷酸激活蛋白激酶間接激活劑的作用機制Figure 3 Activation mechanism of AMPK by indirect activators

3.1.1 5-氨基咪唑-4-甲酰胺核苷 5-氨基咪唑-4-甲酰胺核苷（5-aminoimidazole-4-carboxamide riboside，AICAR，1)，又名阿卡地新（acadesine），是首個被發(fā)現(xiàn)的AMPK激活劑，現(xiàn)已作為研究工具廣泛用于AMPK的相關(guān)實驗研究。AICAR在腺苷轉(zhuǎn)運體的作用下進入細胞，而后在腺苷激酶作用下轉(zhuǎn)化為單磷酸衍生物5-aminoimidazole-4-carboxamide-1-β-D-ribofuranosyl-5'-monophosphate（ZMP）。ZMP與AMP類似，可結(jié)合到AMPKγ亞基，變構(gòu)激活AMPK并阻礙AMPK的去磷酸化，但并不影響細胞內(nèi)AMP/ATP比值[14]。AICAR在2型糖尿病大鼠模型中可降低血漿三酰甘油酯和游離脂肪酸水平、增加機體葡萄糖清除率、減少肝糖原分解、抑制脂肪分解等，從而減輕代謝障礙的諸多癥狀[15]。AICAR在培養(yǎng)的垂體腫瘤細胞內(nèi)可降低p70S6激酶活性，抑制細胞生長[16]，且能在應激條件下引起星形細胞癌凋亡并保護正常腦細胞[17]。但有研究表明，AICAR并不是選擇性AMPK激活劑，其可作用于其他AMP敏感酶，如抑制果糖-1,6-二磷酸酶的活性[18]、刺激糖原磷酸化酶[19]，故在使用AICAR作為AMPK激活劑進行研究時要考慮其可能影響體內(nèi)多種酶活性這一性質(zhì)。

AICAR可抑制核轉(zhuǎn)錄因子-κB（nuclear factor κB，NF-κB）和CCAAT增強子結(jié)合蛋白（CCAAT/enhancerbinding protein，C/EBP）轉(zhuǎn)錄因子進而下調(diào)促炎因子、誘

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Advances in Research on AMPK Activators

XU Yazhou1, LIAO Hong1, ZHANG Luyong1, LI Jia2, PANG Tao1*
(1. New Drug Screening Center, China Pharmaceutical University, Nanjing 210009, China; 2. National Center for Drug Screening, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China)

Adenosine 5′-monophosphate-activated protein kinase (AMPK) is a key player in regulating energy metabolism. It is placed at the center stage in studies of many pathological conditions including metabolic disorders, cardiovascular disease and cancer. The structure and physiological regulation of AMPK have been introduced in this paper. Moreover,the research progresses in indirect and direct activators of AMPK have been especially reviewed, so as to provide insights for the further development of this kind of drugs.

AMPK; indirect activator; direct activator

R962

A

1001-5094（2014）02-0125-07

接受日期：2013-12-16

*通訊作者：龐濤，副研究員；

研究方向：藥物篩選及新藥開發(fā)；

Tel: 025-83271340；E-mail: tpang2012@gmail.com